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| Status |
Public on Dec 22, 2021 |
| Title |
ChIP-Seq WT_Day6_IRF4_10ug |
| Sample type |
SRA |
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| Source name |
P14 CD8+ T cells
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| Organism |
Mus musculus |
| Characteristics |
replicate group: Day6__WT__IRF4
timepoint: WT
tf: IRF4
corresponding input sample for peak calling: April2017_Day6WT_Input
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| Treatment protocol |
On day 6, cells were left untreated or activated with 50 ng/ml PMA and 1 uM ionomycin for 3 hours before harvest for ChIP-seq.
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| Growth protocol |
Naïve P14 CD8+ T cells were isolated with Miltenyi kit (Cat#130-096-543), stimulated with anti-CD3 (2 ug/ml) and anti-CD28 (2 ug/ml) and cultured in RPMI (supplemented with HEPES, NEAA, sodium pyruvate, 2-Mercaptoethanol, 10% FBS) in the presenece of 100 U/ml IL-2.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Briefly, cells were fixed with 1% formaldehyde and genomic DNA was fragmented by Covaris sonicator (E220). The fragmented DNA was then subjected to immunoprecipitation with transcription factor-specific antibodies.
Briefly, the NEBNext ChIP-seq Library Prep Kit (NEB #E6240L) was used to end-repair and adding adaptor to the immunoprecipitated DNA fragments according to the manufacurer's protocol. NEBNext Multiplex Oligos for Illumina Index Primers Sets were then used to PCR amplify and barcode the libraries.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
NextSeq 550 |
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| Description |
WT_Day6_IRF4_10ug
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| Data processing |
All ChIP-seq reads were trimmed using Trimmomatic to remove primer and low-quality bases.
These single-end reads were then aligned to the mm10 reference genome using bowtie2, allowing maximum insert size of 2000 bp, with the “--no-mixed” and “--no-discordant” parameters added.
Reads with a mapping quality (MAPQ) below 30 were removed.
Duplicates were removed with Picard Tools, and the reads mapping to the blacklist regions and mitochondrial DNA were also removed.
Each combination of transcription factor and condition had two replicates, with the exception of Runx3 in WT, which we limited to one high-quality replicate. ChIP-seq peaks were called in each replicate, versus a control sample (listed as "Corresponding Input Sample for Peak Calling" in this dataset) , using MACS2 with a q-value cutoff of 0.05. Overlapping peaks among replicates were merged. A region was considered a valid peak for a transcription factor if it had a q-value below 0.05 in at least two replicates, and a q-value below 0.001 in at least one of the replicates.
Genome_build: mm10
Supplementary_files_format_and_content: Tab-delimited files of peaks identified with macs2, merged across replicates. Peaks considered valid if below qvalue of 0.05 in both replicates and below 0.001 in at least one replicate.
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| Submission date |
Dec 21, 2021 |
| Last update date |
Dec 22, 2021 |
| Contact name |
Jim Kaminski |
| E-mail(s) |
[email protected]
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| Organization name |
Boston Children's Hospital
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| Street address |
1 Blackfan Circle
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
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| Platform ID |
GPL21626 |
| Series (2) |
| GSE192386 |
Batf-mediated Epigenetic Control of Effector CD8+ T Cell Differentiation (ChIP-Seq) |
| GSE192390 |
Batf-mediated Epigenetic Control of Effector CD8+ T Cell Differentiation |
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| Relations |
| BioSample |
SAMN24281767 |
| SRA |
SRX13473862 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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